The primary aim of the proposed research is to continue study of the mechanisms of skin tumor promotion with emphasis on the anthrone class of skin tumor promoters. In addition, we will focus on the role of specific wound-related growth factors, TGFalpha, KGF, and IGF-1 in this process. Furthermore, we will make extensive use of transgenic animals in the next project period. Current evidence indicates that tumor promotion plays an important role in the etiology of human cancer emphasizing the importance of studies related to this process. In addition, it is important to study mechanisms of tumor promotion by compounds other than the phorbol esters since tumor promotion in humans may occur by more than one mechanism or by mechanisms different than that of the phorbol esters. Data obtained during the previous funding period has provided substantial evidence to support the hypothesis that epidermal toxicity plays an important role in tumor promotion by anthrones. This toxicity is due to the oxidation of anthrones and generation of free radical intermediates. Furthermore, we have found that chrysarobin induces a number of biochemical and molecular changes in mouse skin in vivo that mimic responses during would healing including: induction of transforming growth factor alpha (TGFalpha), induction of keratinocyte growth factor (KGF), induction of insulin-like growth factor-1 (IGF-1), and induction of several differentiation markers in a manner similar to their expression during would healing. The phorbol ester, 12-0-tetradecanoylphorbol-13-acetate (TPA), induces similar increases in TGFalpha and differentiation markers both in vivo and in vitro but no effect on KGF or IGF-1 mRNA levels. In the next project period, we will test the hypothesis that anthrones, through induction of epidermal toxicity, promote skin tumor through a coordinated interaction between the signaling pathways mediated by TGFalpha, KGF, and IGF-1. The Specific Aims are: (i) To determine the mechanism for elevated epidermal TGFalpha mRNA and protein synthesis in anthrone- treated skin and its relationship to tumor promotion by this class of compounds; (ii) To examine the role of other EGF receptor ligands (e.g., amphiregulin (AR), heparin-binding EGF-like growth factor (HB-EGF), and betacellulin (BTC) in skin tumor promotion by anthrones; (iii) To examine, in detail, the changes in expression of KGF in skin, potential mechanism(s) for these changes, and to explore further the role of KGF in tumor promotion by anthrones; and (v) To examine in detail the changes in expression of IGF-1, potential mechanisms for these changes, and to explore the importance of IGF-1 in tumor promotion by anthrones.